This invention relates to flow cytometry.
Flow cytometers are frequently used for the analysis of particles such as cells or beads in a number of different applications. The system allows for determination of both particle morphology and evaluation of particle features by detection of optical labels. The ability to distinguish multiple particles sizes and colors allows multiplex application providing higher capacity of this technology to obtain information from analyzed targets.
The word “particle” as used herein means any discrete target that may be optically analyzed, enumerated or sorted by a flow cytometer. This group includes cells, cell fragments, and beads.
A conventional flow cytometry system disclosed in U.S. Pat. Nos. 6,315,952, 6,638,481, and 7,368,084 uses a positive pressure delivery system where a peristaltic pump injects samples into a flow cytometer. The pump is positioned between the microplate sample source and the cytometer, which has two significant consequences. First, it constrains the length of the sample tubing and fluid carryover (transfer of fluid between samples) that is directly proportional to tubing length. With a conventional HyperCyt® peristaltic pump and a sample tubing length of 1 M (0.01″ ID), the fluid carryover between samples of 1-20 is about 10% based on the thin film left behind (J. W. Bartsch et al., Anal. Chem., 76:3810-17, 2004).
A second consequence of positioning a peristaltic pump between a sample source and a flow cytometer is that the passage of cells through the pump exerts compressive forces that can transiently perturb physiology (e.g., intracellular Ca2+ equilibrium). The conventional HyperCyt® high-throughput flow cytometer system must therefore be limited in use to assays insensitive to such physiological perturbations.